This invention Soybean is economically an important crop for feed, oil and other soy products. However, the genetic base of soybean cultivars grown in the United States is extremely narrow. Recent screening of domestic soybean varieties, plant introductions and other accessions at Ft. Detrick Maryland has shown all the varieties tested to have some level of susceptibility to soybean rust isolates recently collected from Asia, Africa, and South American. Most currently-available soybean varieties with sources of resistance to multiple strains of soybean cyst nematode are genetically similar, which indicates that finding new genes for resistance to soybean cyst nematode in domestic soybean germplasm may be difficult.
Soybean rust and cyst nematodes are economically damaging pathogens that threaten soybean production fields within the United States and elsewhere. Soybean rust (SBR) is a foliar soybean disease capable of causing significant economic yield loss (Bromfield, K. R. 1984, “Soybean Rust,” Monogr. 11, American Phytopathological Society, St. Paul, Minn.; Ogle, H. J., D. E. Byth, and R. McLean, 1979, “Effect of rust (Phakopsora pachyrhizi) on soybean yield and quality in southeastern Queensland, Australia,” Aust. J. Agric., Res. 30:883-893; Sinclair, J. B., 1989, “Threats to production in the tropics: Red leaf blotch and leaf rust,” Plant Dis., 73:604-606). In recent years, soybean rust has reduced yields and raised production costs for soybeans in every major production region of the world except the United States. Two fungal species, Phakopsora pachyrhizi Sydow (also known as the Asian species) and P. meibomiae (Arthur) Arthur (also known as the New World species), cause soybean rust and are spread primarily by windborne spores that can be transported over long distances. Asian soybean rust, P. pachyrhizi, the more aggressive of the two species, was first reported in Japan in 1903 and was confined to the Eastern Hemisphere until its presence was documented in Hawaii in the mid-nineties. Soybean rust in the continental United States was first confirmed in November of 2004 (Harmon, P. F., M. T. Momol, J. J. Marois, H. Dankers, and C. L. Harmon, 2005, “Asian soybean rust caused by Phakopsora pachyrhizi on soybean and kudzu in Florida,” Online Plant Health Progress, doi:10.1094/PHP-2005-0613-01-RS).
The number of states with confirmed soybean rust cases in the United States as of the end of 2005 stands at ten. The presence of soybean rust in Louisiana, Mississippi and Florida had been confirmed by USDA's Animal and Plant Health Inspection Service within a week of the initial discovery in Louisiana. Subsequent sampling resulted in confirmations of rust on soybeans in Alabama, Georgia, Arkansas, Missouri, South Carolina, Tennessee, and Texas. The species of soybean rust found on all positive samplings is Asian soybean rust (P. pachyrhizi), the more aggressive of the two species. The timing and distribution of soybean rust growth in the southeastern U.S. is consistent with spores having been brought from South American by Hurricane Ivan.
In the first year of a predicted soybean rust infestation, assuming that U.S. producers are able to treat with fungicides upon soybean rust detection, the expected value of losses across all U.S. agricultural producers and consumers would range from $640 million to $1,341 million, depending on the severity of infestation. U.S. soybean acreage would decline, and soybean prices to consumers would be expected to increase. (Livingston, M. et al. (2004) “Economic and Policy Implications of Wind-Borne Entry of Asian Soybean Rust into the United States,” USDA Outlook Report No. OCS04D02.)
Four major resistance genes that confer specific resistance to Phakopsora pachyrhizi have been identified in Glycine max (Bromfield, K. R. and Hartwig, E. E., 1980, “Resistance to soybean rust and mode of inheritance,” Crop Sci., 20:254-255; Hartwig, E. E. and Bromfield, K. R., 1983, “Relationship among three genes conferring specific resistance to rust in soybean,” Crop Sci., 23:237-239; Hartwig, E. E., 1986, “Identification of a fourth major gene conferring resistance to soybean rust,” Crop Sci., 26:1135-1136). However, isolates of P. pachyrhizi have been identified that are capable of causing a susceptible SBR type of reaction despite the presence of SBR resistance genes (Hartwig, E. E., 1986, “Identification of a fourth major gene conferring resistance to soybean rust,” Crop Sci., 26:1135-1136). No general resistance or tolerance genes of SBR have been identified. The typical SBR resistant response consists of a few lesions that are red-brown in color and typically produce only small amounts of spores and in a fully susceptible reaction, more numerous tan colored lesions appear and produce profuse amounts of spores (Bromfield, K. R., Melching, J. S., and Kinsolver, C. H., 1980, “Virulence and aggressiveness of cultures of Phakopsora pachyrhizi causing soybean rust,” Phytopath., 70:17-21). Application of fungicide is currently the only method of yield protection (Hartman, G. L., Wang, T. C., and Tschanz, A. T., 1991, “Soybean rust development and the quantitative relationship between rust severity and soybean yield,” Plant Dis., 75:596-600).
Nematode infestations are also a significant problem for soybean production. These plant parasites are microscopic worms that cause 1.4 billion dollars in U.S. soybean crop losses annually. Nematodes are found ubiquitously in soybean crops. In most cases, damage caused by cyst nematode is not visually apparent, however, yield loss can still exceed 30% in the absence of foliar symptoms. Seventy percent of soil samples collected in southern Illinois were found to be positive for soybean cyst nematode (SCN, Heterodera glycines Ichinoe), with an average population density of 9,438 eggs per 100 cc of soil. This level of infestation is high enough to cause significant yield loss, even in known resistant varieties (Niblack, T. (2004), “SCN May Be Causing Damage Greater than Expected in Southern Illinois,” University of Illinois Extension Bulletin No. 19 Article 4).
A soybean that is both rust-resistant and nematode-resistant is needed to improve crop yields.
The genus Glycine is divided into two subgenera—Glycine (perennials) and Soja (annuals). The commercial soybean is a member of the subgenus Soja, whose progenitor is the wild annual species G. Soja. The subgenus Glycine contains 26 wild perennial species (Singh, R. J. Nelson, R. L., and Chung, G. H. (2006), Soybean. In R. J. Singh (ed.) Genetic Resources, Chromosome Engineering, and Crop Improvement Oilseed Crops Volume 4. Taylor&Francis Group, Boca Raton, Fla., In press). These species have a wide geographical distribution, and are morphologically, cytologically and genomically diverse. Thus far, soybean breeders have been restricted to the subgenus Soja for improving soybean cultivars and have not exploited the wealth of genetic diversity found in the wild perennial Glycine species.
Wild perennial species of Glycine possess several useful agronomic traits (Burdon, J. J. and Marshal, D. R. (1981), “Inter- and intra-specific diversity in disease response of Glycine species to the leaf rust fungus Phakospora pachyrhizi,” J. Ecol. 69:381-390; Burdon, J. J., 1986, “The potential of Australian native Glycine species as sources of resistance to soybean leaf rust (Phakopsora pachyrhizi),” p. 823-832 In B. Nampompeth and S. Subhadrabandu (eds) “New frontiers in breeding researches,” Faculty of Agriculture, Kasetsart University, Bangkok; Hartman, G. L., Wang, T. C., and Hymowitz, T., 1992, “Sources of resistance to soybean rust in perennial Glycine species,” Plant Dis., 76:396-399), including resistance to a number of fungal soybean diseases (Riggs, R. D., Wang, S., Singh, R. J., and Hymowitz, T., 1998, “Possible transfer of resistance to Heterodera glycines from Glycine tomentella to soybean,” Supp. J. Nematol., 30(4S):547-552; Kenworthy, W. J., 1989, “Potential genetic contributions of wild relatives to soybean improvement,” p. 883-888, In Pascale, A. J. (ed.), World Research Conference IV, Proc., Buenos Aires, 5-9 Mar. 1989, Argentina, Assoc. Soja, Buenos Aires; Hartman, G. L., Wang, T. C., and Hymowitz, T., 1992, “Sources of resistance to soybean rust in perennial Glycine species,” Plant Dis., 76:396-399; Hartman, G. L., Gardner, M. E., Hymowitz, T., and Naidoo, G. C., 2000, “Evaluation of perennial Glycine species for resistance to soybean fungal pathogens that cause sclerotinia stem rot and sudden death syndrome,” Crop Sci., 40:545-549; Schoen, D. J., Burdon, J. J., and Brown, A. H. D., 1992, “Resistance of Glycine tomentella to soybean leaf rust Phakopsora pachyrhizi in relation to ploidy level and geographical distribution,” Theor. App. Genet., 83:827-832) and parasitic nematodes (Riggs, R. D., Wang, S., Singh, R. J., and Hymowitz, T., 1998, “Possible transfer of resistance to Heterodera glycines from Glycine tomentella to soybean,” Supp. J. Nematol., 30(4S):547-552).
Glycine tomentella accessions PI441001 and PI441008 have been found to carry resistance to SBR (Schoen et al. (1992), supra), and SCN (Riggs et al. (1998), supra).
It would be beneficial to introduce these traits into cultivated soybean through wide hybridization. However, there are significant difficulties in utilizing wild perennials as a potential source of SBR resistance as the ploidy level between the two species are not necessarily comparable. A number of sterile intersubgeneric F1 hybrids have been reported (Hymowitz, T. et al. (1998), “The genome of Glycine,” In: Plant Breeding Reviews (ed. J. Janick), John Wiley and Sons, Inc., New York, USA:289-317).
Ladizinsky et al. (Ladizinsky, G., Newell, C. A., and Hymowitz, T. (1979), “Wide crosses in soybeans: prospects and limitations.” Euphytica 28:421-423) attempted to hybridize domestic soybean with five species of wild soybean (G. canescens, G. clandestine, G. falcata, G. tabacina, and G. tomentella). However, they failed to produce viable F1 hybrids.
Singh, R. J. et al. (1990), “Backcross-derived progeny from soybean and Glycine tomentella Hayata intersubgeneric hybrids,” Crop Sci. 30:871-874, reported production, for the first time, of backcross-derived progenies from a synthetic amphiploid (2n=118, genome GGDDEE) of G. max (2n=40, genome GG)×G. tomentella Hayata (PI483218) (2n=78, genome DDEE). (The term “amphidiploid” is also used herein to refer to a plant having an artificially doubled chromosome number.) The objective was to obtain monosomic alien addition lines (MAALs) to study the introgression of genes from G. tomentella responsible for traits such as resistance to SBR, SCN, and tolerance to abiotic stresses (Singh, R. J., et al. (1998), “Monosomic alien addition lines derived from Glycine max (L.) Merr. and G. tomentella Hayata: Production, characterization and breeding behavior,” Crop. Sci. 38:1483-1489). The frequency of seed formation and recovery of plants through immature hybrid embryo rescue was very low and only four pods were obtained from the amphidiploid plants. While backcrossing the amphidiploid (2n=118) with G. max cv. Clark 63 (2n=40), only 15 backcross plants with a chromosome number of 2n=76, instead of the expected chromosome number of 2n=79, were obtained. (Singh, R. J. et al. (1990), “Backcross-derived progeny from soybean and Glycine tomentella Hayata intersubgeneric hybrids,” Crop Sci. 30:871-874.) No other workers have been able to duplicate the production of a fertile hybrid between a wild perennial and domestic soybean.
A summary of efforts to cross wild perennial soybean with domestic soybean to produce hybrids that can be used for further plant breeding is provided in Hymowitz, T. (2004), “Chapter 4, Specification and Cytogenetics” of Soybeans: Improvement, Production, and Uses, 3rd ed., Agronomy Monograph no. 16, American society of Agronomy, Crop Science Society of America, Soil Science Society of America, 677 S. Segoe Rd., Madison, Wis. 53711, USA. The only group which has been able to produce plants capable of breeding with elite domestic soybean lines for the purpose of introgressing useful traits from the wild species into high-yield domestic soybeans, has been that of the inventor.
Broué et al. (1982), “Interspecific hybridization of soybeans and perennial Glycine species indigenous to Australia via embryo culture. Euphytica 31:715-724, disclosed attempts to cross a hybrid between the wild perennial species G. tomentella and G. canescens with domestic G. max. The resulting plant was sterile.
In 1982, two crosses between wild perennial species G. tomentella and domestic G. max were reported, again producing only sterile plants (Newell, C. A. and T. Hymowitz (1982), “Successful wide hybridization between the soybean and a wild perennial relative, G. tomentella Hayata,” Crop Sci. 22:1062-1065).
In 1985, Singh, R. J. and Hymowitz, T. (1985), “Intra- and Interspecific Hybridization in the Genus Glycine Subgenus Glycine Willd.: Chromosome Pairing and Genome Relationships,” Z. Pflanzenzücht. 95(4): 289-310, showed that fertile hybrids could be obtained between certain wild perennial species. Due to the occurrence of pod abortion in distant hybrids, the immature seeds were extracted from the pods and germinated on an artificial medium. However, fertile plants capable of being backcrossed with domestic soybean were not obtained.
Sakai, T. and Kaizuma, N (1985), “Hybrid Embryo Formation in an Intersubgeneric Cross of Soybean (Glycine max Merrill.) with a Wild Relative (Glycine tomentella Hayata),” Japanese Journal of Breeding. 35(4): 363-374, reported production of a hybrid embryo cell in a cross between G. max and G. tomentella. No plant was produced.
Grant, J. E., et al. (1986), “Cytogenetic affinity between the new species Glycine argyrea and its congeners,” Journal of Heredity 77, 423-426, reported crossing a hybrid between the wild species G. argyrea and G. canescens with domestic G. max, with chromosome amplification to produce an amphidiploid that was sterile.
Hood, M. J. and Allen, F. L. (1980), “Interspecific hybridization studies between cultivated soybean, Glycine max and a perennial wild relative, G. falcata,” Agron. Abst. American Society of Agronomy, Madison, Wis. p. 58. and Hood, M. J. and Allen, F. L. (1987), “Crossing Soybeans with a wild perennial relative,” Tennessee Farm and Home Science, 144:26-50 reported attempts to cross G. max and the wild perennial G. falcata, but no hybrid plants were actually obtained (Hymowitz, T., 2004, “Speciation and cytogenetics,” pp. 97-136, In Boerma, H. R. and Specht, J. E. (eds.), “Soybeans: Improvement, production and uses,” 3rd ed., Agron. Monogr. 16 ASA, CSSA, and SSSA, Madison, Wis.). The plants observed were selfed plants rather than hybrids.
Singh, R. J., and Hymowitz, T. (1987), “Intersubgeneric crossability in the genus Glycine Willd.,” Z. Pflanzenzüchtg 98:171-173, reported a sterile cross between domestic G. max and wild G. clandestina. 
Also in 1987, Newell, C. A., et al. (1987), “Interspecific hybrids between the soybean and wild perennial relatives,” J. Hered. 78:301-306, reported the first partially fertile cross between G. tomentella and G. max) after treatment of F1 plants with colchicine. Additional crosses between G. canescens and G. max, and G. tomentella and G. max, after colchicine treatment, were sterile.
Shoemaker, R. C., et al. (1990), “Fertile Progeny of a Hybridization Between Soybean (Glycine max (L.) Merr.) and Glycine tomentella Hayata,” Theoretical and Applied Genetics. 80(1): 17-23 reported that vegetative cuttings from the synthetic amphidiploid obtained from a cross between G. max and G. tomentella reported in Newell et al (1987)., supra, were transferred from the Monsanto Company to Iowa State University during October of 1987. However, this reputed hybrid was examined in a later thesis, Heath, M. S. (1989), “Analysis of Hybrid Plants and Progeny of a Cross Between Glycine max (L.) Merr. and Glycine tomentella,” M. S. Thesis, Iowa State University, and it was discovered that the G. tomentella chromosome complement had been eliminated after genetic exchange and/or genetic modification had taken place between the two genomes. The purported hybridization could not be reproduced by the author or other researchers.
Coble, C. J. and Schapaugh, W. T. (1990), “Nutrient Culture Medium Components Affecting Plant Recovery from Immature Embryos of Three Glycine Genotypes and an Interspecific Hybrid Grown In Vitro,” Euphytica. 50:127-133, reported a sterile G. max×G. tomentella cross.
Singh, R. J. et al. (1990), “Backcross-Derived Progeny from Soybean and Glycine tomentella Hayata Intersubgeneric Hybrids,” Crop Sci. 30:871-874, reported that chromosomes of an F3 hybrid, having 59 chromosomes, were doubled by colchicine treatment and the resulting amphidiploid was pollinated by soybean cultivars Altona, Bonus, Clark 63, Essex, Williams and Wye. The pollinated gynoecia were sprayed 24 hours after pollination with a hormone solution of 100 mg gibberellic acid (GA3), 25 mg 1-naphthalene acetic acid (NAA), and 5 mg kinetin/L distilled water. The F2 plant yielded four pods with two seeds each. One seed was germinated to produce an F3 plant that was multiplied by cloning and grafting onto the soybean. This was then crossed with domestic soybean cultivar Clark 63 to obtain three immature embryos from which 15 plants were regenerated through organogenesis from callus. These plants were pollen and seed sterile, and no second backcross generations were recovered. Five plants were similarly recovered from one callused embryo of the F3 plant crossed with domestic soybean cultivar Essex. These plants contained 98 chromosomes and their use for further backcrossing was not reported.
Chung, G. H., and Kim, J. H. (1990) “Production of interspecific hybrids between Glycine max and G. tomentella through embryo culture,” Euphytica 48:97-101, reported a sterile G. max×G. tomentella cross.
Chang, K. et al. (1991), “Interspecific Hybrids between G. tomentella and Soybean,” Genetic Engineering Res. Inst. Report 6:23-38 (Korea), reported obtaining hybrid embryos; however, no fertile hybrids were reported.
Kim, K. S. (1991), “Obtaining Intersubgeneric Hybridization between Glycine max and G. latifolia,” Plant Tissue Culturing Technology 18(1):3946 (Korea) reported hybridization, but no fertile hybrid plants.
Kwon, C. S. and Change, K. Y. (1991), “In vitro germination of young hybrid embryos between Glycine max and G. tomentella,” Korean J. Breed. 22:379-383, reported production of a sterile hybrid.
Shen, B. H. and Davis, L. C. (1992), “Nodulation and Nodulin Gene Expression in an Interspecific Hybrid between Glycine max and Glycine tomentella,” Australian Journal of Plant Physiology 19(6):693-707 reported hybridization but not fertile hybrids.
Singh, R. J., et al. (1993), “Backcross (BC2-BC4)-Derived Fertile Plants from Glycine max and G. tomentella Intersubgeneric Hybrids,” Crop Sci 33:1002-1007, reported that the amphidiploid described above with respect to their 1990 paper was crossed with Clark 63 to form a first backcross (BC1) plant, which was again crossed with Clark 63 to form a second backcross (BC2) plant, and repeatedly backcrossed with Clark 63 to form BC3-BC6 plants.
Bodanse-Zanettini, M. H. et al. (1996) “Wide hybridization between Brazilian soybean cultivars and wild perennial relatives,” Theoretical and Applied Genetics 93 (5-6):703-709, reported a cross between G. max and G. tomentella with colchicine treatment of the F1 generation to produce an amphidiploid. Attainment of a fertile plant was not reported.
Singh, R. J. et al. (1998), “Monosomic alien addition lines derived from Glycine max (L.) Merr. and G. tomentella Hayata: production, characterization, and breeding behavior,” Crop Sci. 38(6):1483-1489 reported that they had isolated 22 individual monosomic alien addition lines (MAALs) from fertile lines derived from the cross of G. max with G. tomentella. They also reported, in Riggs, R. D., et al. (1998), “Possible Transfer of Resistance to Heterodera glycines from Glycine tomentella to Glycine max,” Supplement to Journal of Nematology 30(4S):547-552, that one of the seeds resulting from BC4 plants reported in Singh, et al. (1993), was germinated and some of these plants were moderately resistant to nematodes of race 3, one of the crosses was resistant to race 5 in a single test, and four were resistant and five moderately resistant to race 14. None were resistant to all three races. However, Brucker, E. A. (2004), “The Effect of the Soybean Gene rhg1 on Reproduction of Heterodera glycines in the Field and Greenhouse and Associated effects on Agronomic Traits,” MS Thesis, University of Illinois at Urbana-Champaign, found no resistance in the backcross generations.
All publications and patent documents referred to herein are incorporated by reference to the extent not inconsistent herewith.